3 [unreadable] [unreadable] COLLABORATING INSTITUTION (S) 3 [unreadable] [unreadable] OVERALL CRITIQUE 3 [unreadable] [unreadable] PROGRESS IN THE CURRENT FUNDING PERIOD 3 [unreadable] [unreadable] PRINCIPAL INVESTIGATOR 4 [unreadable] [unreadable] BUDGETARY OVERLAP 4 [unreadable] [unreadable] HUMAN SUBJECTS RESUME 4 [unreadable] [unreadable] VERTEBRATE ANIMAL RESUME 4 [unreadable] [unreadable] [unreadable] [unreadable] INDIVIDUAL PROJECTS AND CORES [unreadable] [unreadable] [unreadable] Project 2: Pathophysiology of PNH 5 [unreadable] (Robert Brodsky, M.D.) [unreadable] [unreadable] [unreadable] COMMITTEE BUDGET RECOMMENDATIONS [unreadable] [unreadable] REVIEW PANEL ROSTER [unreadable] [unreadable] IRG SUBCOMMITTEE D ROSTER [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] [unreadable] DESCRIPTION (provided by applicant): Hematopoietic Stem Cells for Transplantation. Myelo immunosuppressive therapy followed by blood and marrow transplantation (BMT) offers great promise for cure of most hematologic malignancies, a number of solid cancers, and a variety of fatal nonmalignant disorders. Nevertheless at present, only a minority of patients with diseases potentially curable by BMT are indeed cured, since many are not eligible for BMT and a significant number who are transplanted die of toxicity or disease recurrence. The ability to identify, isolate, and manipulate lymphohematopoietic stem-progenitor cells (HSCs) will improve both the availability and outcome of BMT. [unreadable] [unreadable] The overall goals of this competitive renewal Program Project proposal are to better understand the pathophysiology of normal HSCs and abnormal HSCs in human diseases. Based on this understanding, we propose to develop specific laboratory methods to make HSCs better transplantation tools, for severe aplastic anemia (SPA), PNH, MDS and leukemia, and for BMT in general. Specifically, we propose in this Program Project to: [unreadable] [unreadable] 1. Further improve our novel and highly effective immunoablative but non-myeloablative treatment for SAA (Projects 1, 2), [unreadable] 2. Develop increased understanding of the pathogenesis of SAA and PNH (Projects 1, 2, 3, 4), [unreadable] 3. Investigate the role and regulation of FLT3 in normal and diseased HSCs (Projects 3, 2, 1), and [unreadable] 4. Molecularly describe apoptotic pathway status of T/NK cells mediating graft vs. host disease (GVHD) and SPA, as well as HSCs in SM and PNH; determine if cells transduced to constitutively express the Fas Ligand (FasL) gene functionally protect allogeneic host cells from immune attack in GVHD and autologous HSCs from immune attack in SAAJPNH (Projects 4,2, 1). [unreadable] [unreadable] COLLABORATING INSTITUTION(S): None. [unreadable] [unreadable] OVERALL CRITIQUE: Paroxysmal Nocturnal Hemoglobinuria (PNH) is an uncommon, acquired disorder of blood cells caused by mutation of the phosphatidylinositol glycan class A (PIG-A) gene. The pathophysiology of this disease remains unknown with several hypotheses proposed to explain the "escape" hypothesis whereby the clonal disorder dominates, as well as decreased apoptosis. The basic hypotheses being tested here regarding the pathophysiology of PNH are highly novel and excellent progress has been made during the current funding period. Doubts were raised at the time of the original review whether the investigator would be able to perform the experiments proposed to test his hypothesis, largely based upon the small number of cell available for analysis. In progress to date he has amply demonstrated that he is able to do these experiments and has developed novel approaches to examine these questions. In conclusion, this is a strong project that is highly innovative and the Principal Investigator has answered most of the concerns raised by the original reviewers that limited enthusiasm for the project and that limited funding of the first three years of the program. [unreadable] [unreadable] This project is highly integrated within the program. In particular, there is considerable synergy between this project and Project 1 of the currently funded P01. The unanimous view of the review group was that the infrastructure necessary to accrue and study these patients could only occur within the program project. [unreadable] [unreadable] PROGRESS IN THE CURRENT FUNDING PERIOD: This project was funded for only 3 years at the last review due to concerns that specific techniques in the proposed approach would be feasible. The strong preliminary data included in this application provides evidence that the proposed specific aims have been accomplished in part during these first 3 years and that the new proposed experiments are logical extensions of the data acquired and summarized in the preliminary data section. Excellent progress has been made since the time of the original submission. [unreadable] The new interesting data that has been generated also provide the basis for future, potentially relevant, studies. The progress is also reflected in 3 peer reviewed manuscripts published during 2003-4 by Dr. Brodsky in the field of PNH and Severe Aplastic Anemia (SAA). There is evidence that scientific synergy has occurred as indicated by joint publications with Dr. Jones, preliminary data generated from patient samples obtained from patients enrolled and treated on protocol outlined in Project 1 (R. Jones). New collaboration is identified in preliminary data generated under Specific Aim 2 including mass spectrometry measurements of ceramide performed in collaboration with Dr. Hubbard. [unreadable] [unreadable] PRINCIPAL INVESTIGATOR: Dr. Brodsky is the Project Leader of the project and Dr. Curt Civin is the Principal Investigator of the overall program project. Dr. Brodsky is Assistant Professor of Oncology and Medicine at Johns Hopkins. He is a highly innovative young Investigator and clinical scientist. He obtained his M.D. from Hahnemann University School of Medicine and after a highly productive postdoctoral period at NIH, completed his fellowship at Johns Hopkins University and joined the faculty there. He has been a highly successful translational investigator with an interest in Aplastic anemia, Paroxysmal Nocturnal Hemoglobinuria, myelodysplastic syndrome and autoimmune diseases. He is highly qualified to lead this project. [unreadable] [unreadable] BUDGETARY OVERLAP: None. [unreadable] [unreadable] HUMAN SUBJECTS RESUME: [unreadable] [unreadable] PROTECTION OF HUMAN SUBJECTS (Resume): No concerns. Laboratory studies to be performed in Project 2 will utilize bone marrow specimens from normal donors, PNH patients and MDS patients. Excess mobilized peripheral blood from patients undergoing autologous stem cell transplant will also be used. The research use of this excess that normally would be discarded, does not require IRB approval. The consent for samples to be used is under IRB-approved protocol 00-01-27-09, "Tissue and Cell Procurement Protocol". Proper identification of specimens and maintenance of confidentiality is applied. Potential risks of standard bone marrow sampling and protection against risk are adequately addressed. [unreadable] [unreadable] DATA AND SAFETY MONITORING PLAN: Not applicable. [unreadable] [unreadable] INCLUSION OF WOMEN PLAN (Resume): Acceptable. The inclusion of women is appropriate for the proposed studies. G1A. [unreadable] [unreadable] INCLUSION OF MINORITIES PLAN (Resume): Acceptable. The plan for inclusion of minorities is appropriate. M1A. [unreadable] [unreadable] INCLUSION OF CHILDREN PLAN (Resume): Acceptable. Patients of all ages can be included in these studies. C1A. [unreadable] [unreadable] VERTEBRATE ANIMALS (Resume): None. [unreadable] [unreadable] [unreadable] PROJECT 2: Pathophysiology of PNH [unreadable] (Robert Brodsky, M.D.) [unreadable] [unreadable]